VGSCs are transmembrane proteins responsible for bestowing electrical excitability upon almost all excitable membranes. The pore is gated by depolarization of the cell membrane, transiently allowing Na+ ions to enter into the cell, and generating the upswing of an action potential. Following activation, VGSCs undergo inactivation, limiting the action potential duration, and allowing rapid membrane repolarization is followed by a return to the resting state. All known VGSCs exhibit remarkable functional similarities and this is reflected in a high degree of amino-acid sequence homology. However, natural toxins are known to discriminate well between Na+ channel subtypes. For example, tetrodotoxin (TTX) from the Puffer fish, can selectively block subtypes of neuronal VGSCs at single nanomolar concentrations, whereas other neuronal VGSCs remain unblocked by the toxin at micromolar concentrations. These neuronal VGSCs that are TTX-insensitive or resistant (TTX-R) are found in the peripheral nervous system, and are exclusively associated with nerves involved in the transmission of pain (see e.g. Akopian et al (1999) “The tetrodotoxin-resistant sodium channel SNS plays a specialised role in pain pathways”. Nature Neuroscience 2, 541-548).
WO 97/01577 (University College London) relates to a novel 1,957 amino acid TTX-insensitive VGSC from mammalian sensory neurons (which has been designated Nav 1.8). U.S. Pat. No. 6,184,349 (Syntex) discusses VGSCs. The sodium channel Nav1.8 (also known as SNS or PN3) is expressed exclusively in small diameter sensory neurones that correspond to Aδ or C-fibre nociceptors, which are the cells that transmit pain signals. One key feature of Nav1.8 pharmacology is its resistance to high concentrations of tetrodotoxin (TTX), which blocks most other sodium channels. Evidence for a role of Nav1.8 in pain signalling comes largely from knock out mice and from studies where the channel is downregulated with antisense oligonucleotides. These experiments suggest that Nav1.8 is important in models of inflammatory, neuropathic and visceral pain.
Nav1.9 (SNS2) is also found exclusively in sensory neurones that signal pain and is also resistant to TTX. The properties of the channel suggest that it is not involved in generation or propagation of action potentials but is involved in setting the level of excitability of the cell. There is evidence that G-proteins can activate Nav1.9, which in turn increases neuronal excitability and makes the cell more likely to fire. There is no direct evidence for involvement of Nav1.9 in pain models, but given its function in the cell and the restricted distribution, it could play a major role in producing the is hyper-reactivity associated with many chronic pain states.
Nav 1.3 is found in brains of adult animals and is sensitive to TTX. There is normally no Nav1.3 in sensory neurones, but after nerve damage, levels are upregulated massively. Again there is no direct evidence for involvement of Nav1.3 in pain, but the selective upregulation after nerve injury suggests that it might play a role in transmission of neuropathic pain signals.
p11 is a member of the S-100 family small calcium binding proteins. p11 is also known as annexin-II light chain, lipocortin-II light chain, calpactin I light chain, 42 C, or S-100 related protein, and these terms may be used interchangeably herein. It is present in a variety of cells separately or as a heterotetramer. The heterotetramer is composed of two copies of p36, also known as annexin-II or calpactin-I heavy chain, and two copies of p11. The association of p11 with p36 is Ca2+-independent and of high affinity, and binding of p11 to p36 strongly enhances p36's ability to bundle F-actin. Within the cell, the heterotetramer is localized at the cytoplasmic surface of the plasma membrane in the submembranous cytoskeleton, and it is suggested that the complex may play a role in membrane trafficking events such as exocytosis, endocytosis and cell-cell adhesion. It is also known that p11 is a strong substrate for tyrosine kinase pp60src and its phosphorylation is a negative modulator of tetramerization and protein function.